In our 2004-2009 SPORE, we, studied the prognosfic significance of iriducible nitric oxide synthase (NOS) protein levels in both primary and metastatic melanoma, as well as the geneficand pharmacologic regulafion of its constitutive expression. The results confirmed that iNOS protein is readily detectable in melanoma cell cytoplasm in the majority of patients, and the quanfity, as detected by immunohistochemistry (IHC), provides prognostic information by identifying piatients with poor survival in bPth univariate and mulfivariate analysis (p<0.001), independently of AJCC staging and associated prognosfic criteria. Melanomas are recognized to be heterogeneous based on efiology and somafic mutafional status and have aberrant expression of inflammatory genes and proteins. We aim to expand the tesfing of INOS protein expression as a clinically useful prognosfic marker and propose that INOS repi^esents a node of an identifiable melanoma inflammatory and oxidafive stress network. We further hypothesize that a signature of poor prognosis for melanoma can be gerieratedby directly tesfing tumors for expression of INOS-related inflammatory markers. Testing for associafion of iNOS protein expression and levels with both geriefic alterations (mutations of BRAF and A/RAS) arid mitogen-activated protein kinase (MAPK) pathway acfivafion was also performed in a large series of primary cutaneous tumor biopsies. We report that acfive MAPK can drive iNOS expression and that inhibition of MAPK arid/or 6/?AF inhibits INOS protein expression. iNOS protein in melanoma was also found to be sensifive to down regulation by pharmacologic agents, possibly fpr therapeutic advantage. Pharmacologic inhibifion of INOS or inhibifion of its product, nitric oxide (NO), restores chemosensitivity in iNOS-posifive melanoma cell lines. Building on these results, three new specific aims are now proposed: 1) identify iNOS-related inflammatory nnarker genes expressed in melanoma, as part of a proposed signature 2) determine which candidate marker proteins can be identified in melanoma tumors by standard immunohistochemistry and whether their presence iadds value to the INOS survival prediction model, and 3) test the inflammatory stress pathway as a target for melanonia patient therapy in a Phase l/ll trial using an anti-inflarnmatory drug that inhibits expression of INOS and other inflammatory mediators.